Microglia is myeloid-derived resident cells within the brain but their exact roles in health and disease are still poorly understood. The inability to reliably distinguish microglia from closely related myeloid cells called macrophages confounds many studies of microglial function. Therefore we screened for a new microglial specific gene that would allow the reliable identification, targeting, and characterization of microglia. In our preliminary studies, we identify TM119, a highly expressed transmembrane protein, as a highly expressed microglia-specific marker that is not expressed by macrophages or other peripheral immune cells. In this application, we will develop several new tools based on TM119 expression to selectively identify, isolate, and manipulate microglia. In the first aim, we will develop two antibodies against TM119 for the identification of microglia by immunostaining and isolation of pure microglia from whole brain tissue by immunopanning. In the second aim, we will develop two mouse lines, a TM119/Cre recombinase knock-in mouse and a TM119/CreERT2 BAC transgenic mouse. These mice will drive constitutive (knock-in) or inducible (BAC) Cre expression selectively within microglia. Crossing these mice with other mouse lines that express Cre- dependent genes will allow selective and specific manipulation of microglial genes. In the final aim, we will use the antibody and genetic tools developed in Aims 1 & 2 to acutely purify microglia from adult, developing, or inflamed mouse tissues to generate gene profiles of pure microglia by gene array and RNAseq. These profiles and all tools developed will be made immediately available upon publication to interested researchers. The tools developed in the proposed studies will enable investigators to better understand the roles of microglia, which may prove critical for a better understanding of the pathophysiology and treatment of human neurological diseases. PUBLIC HEALTH RELEVANCE: Microglia is myeloid-derived resident cells within the brain but their exact roles in health and disease are still poorly understood. We have recently identified a highly expressed gene called TMEM119 that is highly expressed specifically by microglia but not macrophages. In this application, we will exploit this discovery to develop new tools to selectively identify, isolate, and manipulate microglia. The tools will be made available to all researchers and will be helpful in elucidating the exact roles of microglia in CNS health and disease. !